Increased synthesis of a multifunctional calcium (Ca2+) binding protein calreticulin (CRT) has been reported under diverse physiologic and pathophysiologic conditions in various tissues including vascular endothelium. We recently reported that angiotensin IV-stimulated increased expression of CRT in lung endothelial cells and the protein-protein interaction between CRT and the endothelial cell isoform of nitric oxide synthase (eNOS) are causally associated with sustained (> 12 hr) activation of eNOS at a post-transcriptional level. The molecular mechanisms of CRT-mediated sustained activation of eNOS appear distinctly unique since all other known post-transcriptional mechanisms involved in activation of eNOS cause only limited or transient activation. At present CRT knows nothing about the molecular interaction and regulation of the catalytic activity of eNOS. Based on our earlier report and preliminary data, we hypothesize that the interaction of CRT with the electron transfer control element (ETE) of eNOS overrides its putative autoinhibitory function resulting in enhanced electron transfer and sustained activation of eNOS, NO/cyclic guanosine 5'-monophosphate (cGMP) production and vasorelaxation response. To test this hypothesis, we will 1) determine whether CRT interaction with ETE in the reductase domain of eNOS is causally associated with increased electron transfer and catalytic activity of eNOS, 2) characterize the molecular interaction between CRT's distinct N-domain, or P-domain (high affinity Ca2+ binding site), or C-domain (low affinity Ca2+ binding site) and ETE responsible for enhanced catalytic activity of eNOS, and 3) verify whether increased expression of CRT or its discrete domains in isolated endothelial cells and in pulmonary artery segments enhances eNOS activity, NO/cGMP production, and vasorelaxation. Identification of a molecular interaction between ETE of eNOS and CRT that causes sustained activation of eNOS will advance our understanding of eNOS regulation for maintaining pulmonary vascular function in context with increased CRT expression under diverse physiologic and pathophysiologic conditions.